The present invention relates generally to apparatus and methods for performing mass spectrometric analyses of material samples and, more particularly, to an improved technique for dissociating parent ions into daughter ions in tandem mass spectrometers.
Mass spectrometry is an analytical technique which relies on the production of ionized fragments from a material sample and subsequent quantification of the fragments based on mass and charge. Typically, positive or negative ions are produced from the sample and accelerated to form an ion beam. Differing mass fractions within the beam are then selected using a mass analyzer, such as a single-focusing or double-focusing magnetic mass analyzer, a time-of-flight mass analyzer, a quadrupole mass analyzer, or the like. A spectrum of fragments having different masses can then be produced, and the identity of compound(s) within the material sample identified based on the spectrum.
An improved form of mass spectrometry, referred to as tandem or MS/MS mass spectrometry has been developed where a mass-selected ion beam (referred to as the parent ion stream) produced by a first mass analyzer is dissociated into a plurality of daughter ion fragments. The daughter ion fragments are then subjected to a second stage mass analyzer, allowing mass quantification of the various daughter ion fractions. Such tandem mass spectrometry has been found to provide much more information on the material being analyzed and to allow for improved discrimination between various species that may be present in a particular sample. Tandem mass spectrometry is discussed in more detail in McLafferty (1981) Science 214:280-287 and Kondrat and Cooks (1978) Anal. Chem. 50:81A-92A.
The present invention is concerned primarily with methods and apparatus for dissociating the parent ion beam into a beam of daughter ions Collision-induced dissociation (CID) is generally employed to reduce the parent ions into the daughter ions. In the predominant technique, the mass-selected parent ions are collided with gas particles, such as helium or hydrogen particles, to convert a portion of the translational energy into internal excitation energy. A number of the excited molecules will then undergo rapid unimolecular dissociation into structurally significant fragment ions, referred to as daughter ions.
The use of a gas to induce collisional dissociation has several drawbacks. First, very small sample sizes, on the order of micrograms, are often too small to provide sufficient parent ions to produce a significant stream of daughter ions. Second, the daughter ions are frequently produced over a very large energy range as a result of the kinetics of the gas collision. Such a large energy spread may necessitate the use of double focusing analyzers to obtain sufficient resolution of the daughter ion spectrum. Even with the best performing tandem equipment, however, the highest practical resolution is usually limited to about 1000 because of the signal loss resulting from the broad energy differential. See, e.g., Johnson and Biemann (1987) Biochem. 26:1209-1214. The introduction of a collision gas can also raise the pressure in the mass spectrometer which can result in poor resolution of high mass, e.g., greater than 1000 d, compounds. See, e.g., Aberth (1986) Anal. Chem. 58:1221-1225. Finally, tandem mass spectrometers using electrostatic energy analyzers often display uncertainty in mass calibration as a result of collision-associated translational energy loss. See, e.g., Bricker and Russell (1986) J. Am. Chem. Soc. 108:6174-6179.
To at least partially overcome these problems, electron-induced dissociation of the molecular species in the parent ion beam has been proposed. In U.S. Pat. No. 4,731,533, the parent ions are directed axially through a quadrupole structure while four sheets of electrons are directed inwardly through the spaces between adjacent pairs of the quadrupole rods. The electron sheets intersect generally along the parent ion beam axis, and collisions between the electrons and the parent molecules result in molecular dissociation. The patent states that such electron-induced collisions result in improved molecular dissociation efficiency and increased reproducibility. The apparatus and method described by the patent, however, suffer from certain deficiencies. In particular, use of the quadrupole structure and four separate electron filament sources renders focusing of the electrons difficult. Moreover, the production and focusing of the electrons is inefficient, requiring relatively large amounts of power to produce a sufficient number of electrons to provide the desired dissociation efficiencies. The high power requirements, in turn, place relatively high heat dissipation requirements on the system which are exacerbated by the presence of the structure in a vacuum. In addition, the long path length of the electron beam and its relatively narrow cross-section will permit space charge repulsion from the electrons to severely limit the electron beam current. Also, the RF field through which the electrons must traverse will shorten the effective duty cycle of the beam and result in the dumping of excessive power by the deflected electrons on the quadrupole rods. Moreover, the need for a tungsten cathode requires using excessive power (on the order of 500 watts) which will make cooling of the system difficult in the vacuum environment of the quadrupole system.
For these reasons, it would be desirable to provide improved apparatus and methods for the electron-induced dissociation of molecular species, particularly of mass-selected parent ion beams in tandem mass spectrometers. In particular, it would be desirable to provide apparatus and methods which achieve high dissociation efficiencies combined with low scattering losses, reduced energy spreading resulting from collisional momentum transfer, and a large transfer of internal energy. It would be further desirable to provide such advantages while utilizing a low power electron source capable of providing an intense electron flux across the path of the ion beam to be dissociated. The apparatus will preferably be of simple and compact construction and will preferably eliminate the need to provide a quadrupole structure for directing the ion beam between the first and second mass analyzers of the tandem mass spectrometer.